Mature red cells express a prominent Ca2+-activated K+ conductance known as the Gardos current, which participates in cell volume regulation. The intermediate-conductance Ca2+- activated K+ channel hIK1 that was recently cloned from a variety of mesenchymal tissues has been proposed as a likely candidate for the "Gardos channel" responsible for this conductance. However, our studies make it probable that more than one type of Ca2+-activated K+ channel exist in erythroid cells. We have found that mRNA encoding the pore-forming subunits of channels derived from two distinct Ca2+-activated K+ channel families ("Intermediate-Conductance," or "IK" Channels, and "Small- Conductance, or "SK" Channels ) is expressed in reticulocytes and in human erythroid cell lines. Furthermore, we have identified three splice variants of the SK channel hSK1, two of which are more highly expressed in erythroid cells than in brain, an organ in which SK channels are especially prominent. One splice variant that accounts for a large proportion of total hSK1 mRNA in reticulocytes has a large deletion in its cytosolic C-terminus that predicts that its gating by calmodulin or other second messengers may be altered. These findings lead us to the view that a complex constellation of Ca2+-activated K+ channels may be required for normal erythroid function and/or development. The specific aims of this proposal are: [1] to discern the structures of the Ca2+-activated K+ channels that we have found in human erythroid cells, [2] to analyze their electrophysiological profiles in cellular expression systems, and [3] to determine whether the C-terminal deletions in the splice variants of hSK1 that we have identified give rise to alterations in calmodulin binding that consequently result in changes in the gating of the channel by Ca2+ ions. The proposed experiments will test the hypotheses: [1] that members of both the "intermediate- conductance" (IK) and "small-conductance" (SK) families of Ca2+- activated K+ channels exist in erythroid cells; and [2] that isoforms of erythroid SK channels with deletions in their cytosolic C-terminal regions exhibit gating profiles different from that of the native channel. Our studies have the potential to inspire new pharmacological strategies to block the Ca2+- activated K+ current of the red cell, which is known to contribute significantly to the morbidity of the sickling disorders through its effect on cell volume.